Semicarbazide sensitive amine oxidase (SSAO)/vascular adhesion protein-1 (VAP-1) is a membrane protein with a dual function. On the one hand, SSAO [EC 1.4.3.6] belongs to the family of copper-containing amine oxidases, its name deriving from its sensitivity to inhibition by a type of carbonyl reagents called semicarbazide (Lyles G A, Int. J. Biochem. Cell. Biol., 1996, 28, 259-274). SSAO catalyzes the oxidative deamination of primary aliphatic and aromatic amines with the following reaction pathway.RNH2+O2+H2O→RCHO+H2O2+NH3 
The enzymatic reaction of the amine results in the formation of the corresponding aldehyde, H2O2 and ammonia; the products formed in the reaction being generally more cytotoxic than the substrates themselves. For the human enzyme, aminoacetone and methylamine have been identified as endogenous physiological substrates.
On the other hand, analyis of the genetic encoding of an adhesion protein revealed the identity of SSAO and human vascular adhesion protein-1 (VAP-1) (Smith D J et al., J. Exp. Med., 1998, 188, 17-27). VAP-1 is a cell adhesion molecule with some special features distinguishing it from other adhesion molecules related to inflammation, such as the monoamine oxidase activity and a restricted expression pattern (Salmi M et al., Science, 1992, 257, 1407-1409; Smith D J et al., J. Exp. Med., 1998, 188, 17-27). The level of VAP-1 is upregulated in the vasculature at sites of inflammation.
Although the substrate specificity of SSAO/VAP-1 partly overlaps with that of monoamine oxidases (MAOs), SSAO/VAP-1 differs from MAO A and MAO B with respect to cofactor (2,4,5-trihydroxy-phenylalanyl quinone (TPQ) for SSAO/VAP-1), biological function, substrates (there are some distinctive substrates), inhibitors and subcellular distribution. Products of SSAO/VAP-1, such as formaldehyde are mainly extracellular. The absence of formaldehyde dehydrogenase from the blood plasma, where SSAO/VAP-1 products are formed, may amplify the potential toxic effects of formaldehyde towards blood vessels.
SSAO/VAP-1 exists as a membrane-bound and as a soluble form in the plasma, its activity displaying a wide tissue distribution. It has been hypothesized that the soluble form is generated via proteolytic cleavage from the membrane-bound form. The major sources of the enzyme are the endothelial cells, smooth muscle cells and adipocytes. Because expression of SSAO is particularly remarkable in the endothelium and the plasma, cytotoxic effects associated with the enzyme may be increased in the highly vascularised tissues, such as the eyes and kidneys, partially explaining late-diabetic complications (Ekblom J. et al., Pharmacol. Res., 1998, 37, 87-92).
SSAO/VAP-1 has a role in the metabolism of biogenic and xenobiotic amines. Products formed in the enzyme reaction (formaldehyde, methylglyoxal and H2O2 for the endogenous substrates) may be involved in processes such as protein cross-linking, formation of advanced-glycation end products or increase of oxidative stress. Higher concentrations of the physiological substrates in diabetes together with the higher enzyme activity observed may lead to a higher production of the cytotoxic agents, therefore may lead to diabetes-associated complications. Treatment of diabetes-associated vasculopathies such as retinopathy, neuropathy and nephropathy with enzyme inhibitors has been proposed.
SSAO/VAP-1 expression is induced during adipogenesis (Fontana E et al., Biochem. J., 2001, 356, 769-777; Moldes M et al., J. Biol. Chem., 1999, 274, 9515-9523), therefore a role for SSAO/VAP-1 in the adipogenic gene program has been suggested. Due to its special features in adipose tissue, SSAO/VAP-1 has been proposed as a potential target for the treatment of obesity (Bour S et al., Biochimie, 2007, 89, 916-925).
SSAO/VAP-1 as an adhesion molecule plays a role in leukocyte trafficking and is involved in an adhesive cascade leading to the transmigration of leukocytes into inflamed tissues from the circulation. In the adhesion cascade both the amine oxidase and the adhesive function of SSAO/VAP-1 take part (Salmi M et al., Immunity, 2001, 14, 265-276), a direct interaction with a leukocyte surface substrate mediating the leukocyte-SSAO/VAP-1 interaction has been proposed. Products of the enzyme reaction of SSAO/VAP-1, such as H2O2, a signalling molecule itself, via the upregulation of other adhesion molecules leading to enhanced leukocyte trafficking may contribute to the escalation of the inflammatory process. Therefore, inhibitors of the enzymatic activity may serve as useful antiinflammatory agents.
SSAO/VAP-1 inhibitors could reduce leukocyte trafficking at sites of inflammation and therefore reduce the inflammatory process as proved by several animal studies (for example: ulcerative colitis—Salter-Cid L M et al., J. Pharm. Exp. Ther., 2005, 315, 553-562; arthritis—Marttila-Ichihara F et al., Arthritis Rheum., 2006, 54, 2852-282862; multiple sclerosis—Wang E Y et al., J. Med. Chem., 2006, 49, 2166-2173; uveitis—Noda K et al., FASEB J., 2008, 22, 1094-1103). As translocation of VAP-1 to the endothelial cell surface occurs at sites of inflammation, modulation of the normal immune system could be avoided by the use of SSAO/VAP-1 as a novel anti-inflammatory target.
In healthy humans, the plasma SSAO/VAP-1 activity is rather constant. Elevated SSAO/VAP-1 levels or overexpression of the enzyme have been observed in various pathological conditions or diseases, such as diabetes (both type I and type II), particularly in the presence of diabetic complications (Boomsma F et al., Biochim. Biophys. Acta, 2003, 1647, 48-54; Boomsma F, Clin. Sci., 1995, 88, 675-679; Garpenstrand H et al., Diabetic. Med., 1999, 16, 514-521; Meszaros Z et al., Metab. Clin. Exp., 1999, 48, 113-117; Boomsma F et al., Diabetologia, 1999, 42, 233-237; Salmi M et al., Am. J. Pathol., 2002, 161, 2255-2262), congestive heart failure (Boomsma F et al., Cardiovasc. Res., 1997, 33, 387-391), obesity (Meszaros Z et al., Metab. Clin. Exp., 1999, 48, 113-117; Weiss H G et al., Metab. Clin. Exp., 2003, 52, 688-692), end-stage renal disease (Kurkijarvi R et al., Eur. J. Immunol., 2001, 31, 2876-2884), multiple sclerosis (Airas L et al., J. Neuroimmunol., 2006, 177, 132-135), inflammatory liver diseases (Kurkijarvi R et al., J. Immunol., 1998, 161, 1549-1557), psoriasis (Madej A et al., J. Eur. Acad. Dermatol. Venereol., 2007, 21, 72-78), Alzheimer's disease (del Mar Hernandez M et al., Neurosci. Lett., 2005, 384, 183-187; Ferrer I et al., Neurosci. Lett., 2002, 321, 21-24) and myopathies (Olive M et al., Muscle Nerve, 2004, 29, 261-266). A role for SSAO/VAP-1 in apoptosis, possibly leading to vascular tissue damage and atherogenesis has been implicated.
An oxime prodrug approach for ketone drugs, the non-steroidal antiinflammatory drugs ketoprofen and nabumetone, has been reported recently (Kumpulainen H, J. Med. Chem., 2006, 49, 1207-1211). The oxime structure was activated to the ketone with simultaneous release of nitric oxide (NO).
Because of its proposed involvement in a number of inflammatory processes and various pathologies, there is a great demand for inhibitors of SSAO/VAP-1 that can have therapeutic value in the prevention or the treatment of disorders or diseases associated with an elevated level or overexpression of SSAO/VAP-1, said diseases involving acute and chronic inflammations, diseases related to carbohydrate metabolism, diabetes-associated complications, diabetic retinopathy and macular oedema, diseases related to adipocyte or smooth muscle dysfunctions, neurodegenerative diseases and vascular diseases.
Several small-molecule inhibitors of SSAO/VAP-1 have been identified: hydrazine derivatives, phenylallylhydrazines (WO2006/094201, WO2005/014530), hydrazino alcohols and hydrazino indanes (WO2002/0202090, WO2003/006003, WO2005/080319), arylalkylamines, propenyl- and propargylamines, oxazolidinones, haloalkylamines, 1,3,4-oxadiazines (WO2002/0202541), 4,5,6,7-tetrahydroimidazo [4,5-c] pyridines (WO2002/0238153), thiocarbamoyl derivatives, carboxamides and sulfonamides (WO2006/013209, US2007/066646), thiazole derivatives (WO2004/087138, WO2004/067521, WO2006/028269, WO2006/011631), compounds disclosed in WO2005/082343; (compounds reviewed in: Matyus P et al., Curr. Med. Chem., 2004, 11, 1285-1298; Dunkel P et al., Curr. Med. Chem., 2008, 15, 1827-1839). Further relevant documents are mentioned in “Disclosure of the Invention” part below.
We now found that SSAO inhibitors, in particular a special class of compounds containing an oxime group and an unsaturated ring system joining to the carbon atom of the oxime group, optionally through an alkylene moiety, and 2-phenyl-2-propen-1-yl)hydrazine [2-phenylallylhydrazine; see e.g. in Tetrahedron Letters, 36, 3155-3158 (1977), WO2006/094201 and WO2005/014530] exhibit potent analgesic effects in traumatic neuropathy and neurogenic inflammation.
These pathophysiological conditions are mediated by the mechanical damage of peptidergic sensory nerves (in case of traumatic neuropathy) or chemical activation of peptidergic sensory nerves (in case of neurogenic inflammation), resulting in pathological activation and dysfunctions of peptidergic sensory nerves. The undesired activation and dysfunction is implicated in severe hyperalgesia (the threshold of a painful stimulus causing nocifensive behaviour remarkably decreases) and allodynia (non-painful stimulus becomes painful and induces nocifensive behaviour).
Traumatic neuropathy induced by mechanical nerve damage (e.g. suffered in accidents, bone fractures or operations) is mediated by complex mechanisms at the levels of both the peripheral and central nerve endings in the respective innervated region, spinal dorsal horn and other pain-related brain regions. These are completely different mechanisms than the ones involved in metabolic (diabetic) and toxic polyneuropathic conditions, therefore, the present invention may by no means be considered to be derived and extrapolated from the currently documented relations of SSAO and diabetic complications. Metabolic neuropathies (caused by diabetes, uremia etc.) and toxic neuropathies (caused by chemotherapeutic agents, alcohol etc.) are polyneuropathies affecting the nerves throughout the body. They are primarily due to severe biochemical abnormalities within the neurons. In case of diabetic neuropathy, these pathological changes are the consequences of hyperglycemia resulting in the damage of blood vessels around the nerve fibres and also detrimental changes in the metabolic state of the neurons (Kles K K and Vinik A I, Curr. Diab. Rev., 2006, 2, 131-145). These abnormalities lead to the production of free radicals and activation of some signal transduction pathways inducing neural dysfunctions (Ceriello A, Diabetes Care, 2003, 26, 1589-1596).
However, neuropathies caused by traumatic events (mechanical damage-induced axonopathies) affect only one or a few anatomical structures (mononeuropathy) and result in pathological activation and dysfunctions of peptidergic sensory nerves. In these cases, neuropathic pain is caused by different mechanisms compared to metabolic or toxic polyneuropathies, such as abnormal crosstalk between sensory and sympathetic nerves, changes in the expression of different ion channels, marked glial cell activation etc., and mediated by different signaling molecules than in diabetic neuropathy (Banoliel R et al., Oral Dis., 2012, 18(4):317-32; Aley K O. and Levine J D, Neuroscience., 2002, 111(2):389-97). There are also differences in the therapy of diabetic and traumatic neuropathies. Causal pharmacotherapeutic agents (alpha-lipoic acid, benfotiamine) exerting an action based on the pathophysiological mechanism of the disease is only available in diabetic, but not in traumatic neuropathy (Miranda-Massari J R et al., Curr Clin Pharmacol., 2011, 6(4):260-73). Drugs used as symptomatic therapy for diabetic neuropathic pain include antidepressants, anticonvulsants, opioids and some other groups (e.g.: topical lidocain, capsaicin). Their effectiveness is well documented by clinical evidence-based data in painful polyneuropathies (such as diabetic) and postherpetic neuralgia, but not in traumatic neuropathy. Additionally, these drugs do not treat the cause of neuropathic pain and are not effective in a large proportion of patients (Finnerup et al., Pain., 2010, 150(3):573-81).
Peripheral and central sensitization mechanisms play important roles in the development of severe persistent chronic pain induced by mechanical nerve damage, which is not effectively treated by the conventional analgesics. Therefore, intensive traumatic neuropathic pain is a clinically challenging problem, since opioids and cyclo-oxygenase (COX) inhibitor non-steroidal anti-inflammatory agents (NSAIDs) are not potent in these conditions. Adjuvant analgesics, such as certain antiepileptics and antidepressants acting in the central nervous system (CNS) by inhibiting the ascending pain pathway and/or activating the descending inhibitory pathway can be used in some patients, but they cannot be regarded as optimal therapeutic solutions for the problems due to severe acute side effects (cardiovasular, CNS) and/or chronic toxicity.
Furthermore, neurogenic inflammation (vasodilatation, plasma protein extravasation, inflammatory cell activation) induced by the stimulation of peptidergic sensory nerves and the released pro-inflammatory sensory neuropeptides (substance P, calcitonin gene-related peptide) play a very important role in a variety of different acute and chronic inflammatory painful diseases (Chiu et al., Nat. Neurosci., 2013, 15(8):1063-7.), although it is not the exclusive mechanism. This is a basically different inflammatory mechanism compared to immune cell-mediated processes, it is often the very early initiation step even in chronic diseases, which triggers and then remarkably augments further cellular pathways. The neurogenic inflammatory component is not inhibited by the conventional anti-inflammatory drugs (COX inhibitors), and glucocorticoids are only moderately effective in extremely high doses in which they exert very many severe side-effects that limit their clinical applications. Therefore, it is particularly important to identify novel therapeutical mechanisms and targets to inhibit neurogenic inflammatory pain. This could substantially help the treatment of chronic inflammatory disorders providing long-term therapeutical benefits for a great patient population.